The ability to deliver pathogen-resistance genes into mosquito populations has long been sought as a potential alternative for disrupting dengue or malaria transmission where funds and infrastructure are the limiting factors in effective mosquito control. The recent development of effective gene drive transgenes based on CRISPR/Cas9 has largely solved the technical challenges of achieving super-Mendelian introgression, however there exists no means to control or recall such genetic elements once released making safety testing in the relevant environments problematic. Drosophila melanogaster is an extremely trackable genetic model organism, while Aedes aegypti is the main vector of dengue, yellow fever and chikungunya viruses, as well as a model system for studies of other mosquitoes. In this project, we will employ both D. melanogaster and A. aegypti to evaluate a transgene self-elimination strategy whereby a transgene can be pre-programmed to first drive itself into a population and then remove itself from the population without any intervention from the experimenter. We will characterize some of the limiting parameters of the self-elimination system, such as direct repeat length (Aim 1) and number and type of nuclease targeting sites (Aim 2). Finally, we will test this self-elimination strategy in the context of an active gene drive in both flies and mosquitoes (Aim 3). Our innovative approach takes advantage of naturally occurring processes that are conserved throughout eukaryota to completely eliminate all transgenic sequences following potential field releases. Thus, we anticipate that this project will dramatically alter the National and International conversations concerning gene drive technology as a whole, and will raise expectations for what is possible in any future trial to generate pathogen-resistant mosquitoes.